In order to characterize the transforming gene of the McDonough (SM) strain of FeSV, two approaches were undertaken to isolate the SM-FeSV viral DNA. Unintegrated viral DNA was obtained from newly infected cells and analyzed by agarose gel electrophoresis, blotting, and molecular hybridization with appropriate cDNA probes. Major linear forms of 9.5 and 6.5 kbp, respectively, presumably represented helper and sarcoma viral DNA. A preliminary restriction endonuclease map of the unintegrated form of SM-FeSV DNA has been obtained. Restriction endonuclease and Southern blotting techniques have also been utilized to identify viral DNA fragments in the DNAs of SM-FeSV transformed NRK and NIH 3T3 cells. Appropriate sized fragments for molecular cloning have been identified. Following molecular cloning of the SM-FeSV genome, a more detailed restriction endonuclease map will be obtained and the transforming region of the genome will be mapped by analysis of transfection by subgenomic DNA fragments.